Boundary-sensing mechanism in branched microtubule networks

The self-organization of cytoskeletal biopolymers, such as microtubules (MTs), depends on mechanosensing and adaptation to confined spaces such as cellular protrusions. Understanding how these active biopolymers coordinate their formation under confinement leads to advances in bioengineering. Here we report the self-organization of branched MT networks in channels with narrow junctions and closed ends, mimicking cellular protrusions. We find that branching MT nucleation occurs in the post-narrowing region only if this region exceeds a minimum length, determined by MT dynamic instability at the closed end and the timescale for nucleation at a distant point. We term this feedback ‘boundary sensing’. Increasing the amount of branching factor TPX2 in the system accelerates MT nucleation and adjusts this minimum length, but excess TPX2 stabilizes MTs at the closed end, disrupting network formation. We performed experiments and simulations to study how this tunable feedback, wherein growing MTs navigate confinement and create nucleation sites, shapes MT architecture. Our findings impact the understanding of MT self-organization during axonal growth, dendrite formation, plant development, fungal guidance and the engineering of biomaterials. Uncovering the rules of microtubule network self-organization under confinement is key to understanding how cells build structure in complex environments. This study reveals a tunable boundary-sensing feedback mechanism, wherein pioneer microtubules navigate confined environments and generate nucleation sites for new microtubules, thereby shaping network architecture.